United States Prevention, Pesticides EPA 738-F-01-013 Environmental Protection and Toxic Substances September 2001 Agency (7508C)

Acephate Facts

EPA has assessed the risks of acephate and reached an Interim Reregistration Eligibility

Decision (IRED) for this organophosphate (OP) pesticide. Provided that risk mitigation measures are adopted, acephate fits into its own “risk cup”-- its individual, aggregate risks are within acceptable levels. Acephate also is eligible for reregistration, pending a full reassessment of the cumulative risk from all OPs.

Acephate residues in food and drinking water do not pose risk concerns, and by reducing exposure in homes and through residential lawns, acephate fits into its own “risk cup.” EPA made this determination after the registrants agreed to drop indoor residential uses and certain turf uses. With other mitigation measures, acephate’s worker and ecological risks also will be below levels of concern for reregistration.

EPA’s next step under the Food Quality Protection Act (FQPA) is to consider risks from cumulative exposure to all the OP pesticides, which share a common mechanism of toxicity. The interim decision on acephate cannot be considered final until the cumulative risk has been considered. Further risk mitigation may be warranted at that time.

EPA is reviewing the OP pesticides to determine whether they meet current health and safety standards. Older OPs need decisions about their eligibility for reregistration under FIFRA. OPs with residues in food, drinking water, and other non-occupational exposures also must be reassessed to make sure they meet the new FQPA safety standard.

The OP Pilot Public Participation Process

The organophosphates are a group of related pesticides that affect the functioning of the nervous system. They are among EPA’s highest priority for review under the Food Quality Protection Act.

EPA is encouraging the public to participate in the review of the OP pesticides. Through a six-phased pilot public participation process, the Agency is releasing for review and comment its preliminary and revised scientific risk assessments for individual OPs. (Please contact the OP Docket, telephone 703-305-5805, or see EPA’s web site, www.epa.gov/pesticides/op .)

EPA is exchanging information with stakeholders and the public about the OPs, their uses, and risks through Technical Briefings, stakeholder meetings, and other fora. USDA is coordinating input from growers and other OP pesticide users.

Based on current information from interested stakeholders and the public, EPA is making interim risk management decisions for individual OP pesticides, and will make final decisions after the cumulative risk from all OPs has been considered.

The acephate interim decision was made through the OP pilot public participation process, a process that increases transparency and maximizes stakeholder involvement in EPA’s development of risk assessments and risk management decisions. EPA worked extensively with affected parties to reach the decisions presented in this interim decision document that concludes the OP pilot process for acephate.

Uses

• Acephate is an organophosphate insecticide currently registered for use on a variety of field, fruit, and vegetable crops (e.g., cotton, tobacco, cranberries, mint); in food handling establishments; on ornamental plants both in greenhouses and outdoors (e.g., nonbearing fruit trees, Christmas trees, and cut flowers); and in and around the home.

• Annual domestic use is approximately 4 to 5 million pounds of active ingredient per year.

Health Effects

• Acephate can cause cholinesterase inhibition in humans; that is, it can overstimulate the nervous system causing nausea, dizziness, confusion, and at very high exposures (e.g., accidents or major spills), respiratory paralysis and death.

Risks

• Dietary exposures to acephate from eating food crops treated with acephate are below the level of concern for the entire U.S. population, including infants and children. Drinking water is not a significant source of acephate exposure. However, people in the U.S. may be exposed to amounts of the acephate degradate methamidophos through food and drinking water as a result of acephate use. This exposure will be more fully addressed in the methamidophos IRED.

• EPA found risks are of concern for homeowners and children entering homes and lawn areas treated with acephate (excluding golf courses and spot or mound treatments for ant control).

• For agricultural and turf/Pest Control Operator (PCO) uses of acephate, several mixer/loader/applicator risk scenarios currently exceed the Agency’s level of concern. In addition, there are postapplication risks from the use of acephate in cut flowers.

• Ecological risks are also of concern to the Agency. Acephate and its degradate methamidophos are highly toxic to honey bees and beneficial predatory insects on an acute contact basis. Acute and chronic risks to birds and chronic risk to mammals are also high.

2

Risk Mitigation

Dietary Risk

No mitigation is necessary at this time for any dietary exposure to acephate. The acute and chronic dietary risks from acephate do not exceed the Agency’s level of concern.

However, the Agency reserves the right to require further acephate mitigation to address risks from methamidophos residues resulting from acephate uses. Any additional mitigation measures will be addressed when the methamidophos interim RED is completed.

Occupational Risk

In order to mitigate occupational risks, the following risk mitigation measures are necessary:

• Formulate all soluble powder formulations into water soluble bags, except for soluble powders sold for fire ant, harvester ant, or hopper box seed treatment uses.

• Limit the 1 pound active ingredient per acre (lb ai/A) cotton aerial application rate to cotton grown in California and Arizona; reduce the maximum aerial application rate for cotton to 0.75 ai/A for all other areas of the United States.

• Delete aerial application to turf. • Require enclosed cockpits and mechanical flagging for all aerial applications. • Reduce maximum sod farm and golf course turf application rates (non-granular formulations)

to 3 lb ai/A and 4 lb ai/A, respectively. • Reduce maximum application rates for greenhouse floral and foliage plant crops, and outdoor

floral and ground covers to 1 lb ai per 100 gallons water (not to exceed 0.75 lb ai/A for cut flowers and 1.0 lb ai/A for other ornamentals).

• Delete the application of acephate by low pressure handwand to treat trees, shrubs, and outdoor flora; for the control of wasps; and for perimeter treatment by PCOs.

• Delete the use of granular formulations to be applied by belly grinder, shaker can, or by hand to trees, shrubs, and 12" pots.

• Add personal protective equipment to end use product labels for workers who mix and load, and/or apply acephate.

Residential Risk

In order to mitigate residential postapplication risk, the following risk mitigation measures are necessary:

• Delete residential indoor uses. • Delete all turfgrass uses (except golf course, sod farm, and spot or mound treatment for ant

control). • Establish a 3 day pre-harvest interval (PHI) for the harvesting of sod.

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Ecological Risk

The Agency has determined that the following mitigation measures are needed to address ecological risk concerns:

• Establish minimum spray intervals for all agricultural crops of 3 days for application rates up to 0.5 lb ai/A and of 7 days for application rates greater than 0.5 lb ai/A.

• Require labeling to protect honeybees. • Require labeling to reduce the potential for spray drift.

In addition, the measures to reduce occupational and residential risk will also reduce environmental loading and the potential impact to non-target organisms.

Next Steps

• Numerous opportunities for public comment were offered as this decision was being developed. The acephate IRED therefore is issued in final (see www.epa.gov/pesticides/reregistration/status.htm or www.epa.gov/pesticides/op ), without a formal public comment period. The docket remains open, however, and any comments submitted in the future will be placed in this public docket.

• In addition, further mitigation of acephate uses may be necessary to reduce risks from methamidophos residues that result from acephate applications. Once the methamidophos IRED is complete, the Agency will determine whether the methamidophos exposure resulting from acephate use poses risk concerns. Any potential further mitigation will be discussed at the time the methamidophos interim RED is released.

• When the cumulative risk assessment for all organophosphate pesticides is completed, EPA will issue its final tolerance reassessment decision for acephate and may request further risk mitigation measures. The Agency will revoke 3 tolerances and lower 4 tolerances for acephate now. Reassessment of 14 tolerances will be made once additional residue data on cotton gin byproducts have been reviewed. For all OPs, raising and/or establishing tolerances will be considered once a cumulative assessment is completed.

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EUROPEAN COMMISSION HEALTH & CONSUMER PROTECTION DIRECTORATE-GENERAL Directorate E – Food Safety: plant health, animal health and welfare, international questions E1 - Plant health

Acephate

SANCO/3057/99-final

31 May 2002

Review report for the active substance acephate

Finalised in the Standing Committee on the Food Chain and Animal Health at its meeting on 18 October 2002 in support of a decision concerning the non-inclusion of acephate in Annex I

of Directive 91/414/EEC and the withdrawal of authorisations for plant protection products containing this active substance

1. Procedure followed for the re-evaluation process This review report has been established as a result of the re-evaluation of acephate, made in the context of the work programme for review of existing active substances provided for in Article 8(2) of Directive 91/414/EEC concerning the placing of plant protection products on the market, with a view to the possible inclusion of this substance in Annex I to the Directive. Commission Regulation (EEC) No 3600/92(1) laying down the detailed rules for the implementation of the first stage of the programme of work referred to in Article 8(2) of Council Directive 91/414/EEC, as last amended by Regulation (EC) No 1972/99(2), has laid down the detailed rules on the procedure according to which the re-evaluation has to be carried out. Acephate is one of the 90 existing active substances covered by this Regulation. In accordance with the provisions of Article 4 of Regulation (EEC) No 3600/92, United Phosphorus Ltd. on 26 July 1993, K & N EFTHYMIADIS SA on 19 July 1993, Cequisa on 23 July 1993, SANC on 23 July 1993, Tomen France SA on 22 July 1993, Iberotam on 26 July 1993, Industrias Quimicas del Valles on 28 July 1993, Pilar Iberica SL Juan Amich Gali on 23 July 1993, Helm AG on 23 July 1993, Industrias Afrasa on 27 July 1993, B.V. Luxan Registration Department on 21 July 1993 and B& S Buy & Sell GmbH on 28 April 1995 notified to the Commission of their wish to secure the inclusion of the active substance acephate in Annex I to the Directive. In accordance with the provisions of Article 5 of Regulation (EEC) No 3600/92, the Commission, by its Regulation (EEC) No 933/94(3), as last amended by Regulation (EC) No 2230/95(4), designated Italy as rapporteur Member State to carry out the assessment of acephate

1 OJ No L 366, 15.12.1992, p.10. 2 OJ No L 244, 16.09.1999, p.41. 3 OJ No L 107, 28.04.1994, p.8. 4 OJ No L 225, 22.09.1995, p.1.

- 2 -

on the basis of the dossiers submitted by the notifiers. In the same Regulation, the Commission specified furthermore the deadline for the notifiers with regard to the submission to the rapporteur Member States of the dossiers required under Article 6(2) of Regulation (EEC) No 3600/92, as well as for other parties with regard to further technical and scientific information; for acephate this deadline was 30 April 1995. Tomen France S.A., Sinon EU Corporation, M/s Cequisa and B & S Buy and Sell GmbH submitted each a dossier to the rapporteur Member State. Tomen France S.A. was the main data submitter, with a dossier which did not contain substantial data gaps, taking into account the supported uses. Sinon EU Corporation, M/s Cequisa and B & S Buy and Sell GmbH did not submit complete dossiers. In accordance with the provisions of Article 7(1) of Regulation (EEC) No 3600/92, Italy submitted on 30 September 1996 to the Commission the report of its examination, hereafter referred to as the draft assessment report, including, as required, a recommendation concerning the possible inclusion of acephate in Annex I to the Directive. Moreover, in accordance with the same provisions, the Commission and the Member States received also the summary dossier on acephate from Tomen France S.A., on 24 October 1996. In accordance with the provisions of Article 7(3) of Regulation (EEC) No 3600/92, the Commission forwarded for consultation the draft assessment report to all the Member States as well as to Tomen France S.A. being the main data submitter, on 30 August 1999, 8 September 1999. The Commission organised an intensive consultation of technical experts from a certain number of Member States, to review the draft assessment report and the comments received thereon (peer review), in particular on each of the following disciplines: - identity and physical /chemical properties ; - fate and behaviour in the environment ; - ecotoxicology ; - mammalian toxicology ; - residues and analytical methods ; - regulatory questions. The meetings for this consultation were organised on behalf of the Commission by the Biologische Bundesanstalt für Land und Forstwirtschaft (BBA) in Braunschweig, Germany, from November 1999 to July 2000. The report of the peer review (i.e. full report) was circulated, for further consultation, to Member States and the main data submitter on 15 June 2001 for comments and further clarification. In accordance with the provisions of Article 6(4) of Directive 91/414/EEC concerning consultation in the light of a possible unfavourable decision for the active substance the Commission organised a tripartite meeting with the main data submitter and the rapporteur Member State for this active substance on 18 June 2002. In accordance with the provisions of Article 7(3) of Regulation (EEC) No 3600/92, the dossier, the draft assessment report, the peer review report (i.e. full report) and the comments and

- 3 -

clarifications on the remaining issues, received after the peer review were referred to the Standing Committee on the Food Chain and Animal Health , and specialised working groups of this Committee, for final examination, with participation of experts from the 15 Member States. This final examination took place from February 2001 to June 2002, and was finalised in the meeting of the Standing Committee on 18 October 2002. The present review report contains the conclusions of this final examination; given the importance of the draft assessment report, the peer review report (i.e. full report) and the comments and clarifications submitted after the peer review as basic information for the final examination process, these documents are considered respectively as background documents A, B and C to this review report and are part of it. 2. Purposes of this review report This review report including the background documents has been developed and finalised in support of Decision 03/219/EC5 concerning the non-inclusion (at this stage) of acephate in Annex I to Directive 91/414/EEC. In accordance with the provisions of Article 7(6) of Regulation (EEC) No 3600/92, Member States will keep available or make available this review report for consultation by any interested parties or will make it available to them on their specific request. Moreover the Commission will send a copy of this review report (not including the background documents) to all operators having notified for this active substance under Article 4(1) of this Regulation. 3. Overall conclusion in the context of Directive 91/414/EEC The overall conclusion of this evaluation, based on the information available and the proposed conditions of use, is that: - the information available is insufficient to satisfy the requirements set out in Annex II

and Annex III Directive 91/414/EEC in particular with regard to acute consumer exposure and non-target organisms

- concerns were identified with regard to

� acute consumer exposure � non-target organisms, in particular non-target arthropods, birds and mammals, as

well as aquatic organisms

In conclusion from the assessments made on the basis of the submitted information, no plant protection products containing the active substance concerned is expected to satisfy in general the requirements laid down in Article 5 (1) (a) and (b) of Council Directive 91/414/EEC.

5 OJ N° L82, 29.3.03, p. 40.

Article i

Persistence and residual toxicity of some insecticides against Phenacoccussolenopsis on cotton

(Gossypiumspp)

Article in Indian Journal of Agricultural Sciences 79(3):203-206 · March 2009 with 57 Reads

Cite this publication

Ashok Dhawan

20.29Punjab Agricultural University

Sushil Saini

44.42Independent Researcher

Kamaldeep Singh

30.92Alliance University

AnandAneja

Abstract

The persistence and residual toxicity of profenophos, thiodicarb, buprofezin, quinalphos,

chlorantraniliprole, spirotetremate, acephate, carbaryl and chlorpyriphos at recommended

concentrations in field on cotton (Gossypiumspp) against first instar and adult female of mealy bug

Phenacoccussolenopsis Tinsley. The mortality data for the crawlers were taken up to 24 hr, whereas

for the adult females, the mortality data were recorded up to 72 hr within different exposure

periods of 1,3,7,10 and 14 days after application of the insecticides. Based on the index of

persistence toxicity, the order of effectiveness for the crawlers of P. solenopsis were profenophos

(637.1) >thiodicarb (587.9) >buprofezin (492.3) > quinalphos (407.9) >chlorantraniliprole (380.0)

>spirotetramate (371.6) >acephate (345.7) >carbaryl (314.6) >chlorpyriphos (273.4). Similarly, for

the adult female this order of effectiveness was profenophos (572.9) >buprofezin (483.1) >thiodicarb

(430.0) > quinalphos (379.9) >chlorantraniliprole (371.6) >spirotetramate (361.3) >acephate (296.7)

>carbaryl (260.0) >chlorpyriphos (252.3).

Sonoda S, Igaki C. Characterization of acephate resistance in the diamondback moth

Plutellaxylostella. Pesticide Biochemistry and Physiology

Article in Pesticide Biochemistry and Physiology 98(1):121-127 · September 2010 with 32 Reads

DOI: 10.1016/j.pestbp.2010.05.010

Cite this publication

Shoji Sonoda

Chikako Igaki

Abstract

Decreased acetylcholinesterase (AChE) sensitivity and metabolic detoxification mediated by

glutathione S-transferases (GSTs) were examined for their involvement in resistance to acephatein

the diamondback moth, Plutellaxylostella. The resistant strain showed 47.5-fold higher acephate

resistance than the susceptible strain had. However, the resistant strain was only 2.3-fold more

resistant to prothiofos than the susceptible strain. The resistant strain included insects having the

A298S and G324A mutations in AChE1, which are reportedly involved in prothiofos resistance in P.

xylostella, showing reduced AChE sensitivity to inhibition by methamidophos, suggesting that

decreased AChE1 sensitivity is one factor conferring acephate resistance. However, allele

frequencies at both mutation sites in the resistant strain were low (only 26%). These results suggest

that other factors such as GSTs are involved in acephate resistance. Expression of GST genes

available in P. xylostella to date was examined using the resistant and susceptible strains, revealing

no significant correlation between the expression and resistance levels.

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Outline

Abstract

Keywords

1. Introduction

2. Materials and methods

3. Results

4. Discussion

Acknowledgments

References

Figures (4)

Fig. 1. Frequencies of the A298S and G324A mutations in the SS and MR strains of…

Fig. 2. Inhibitory effect of methamidophos on crude AChE preparations from SS and MR…

Fig. 3. Amino acid sequence alignment of the delta GSTs from Plutellaxylostella (Pxd),…

Fig. 4. Quantification of expression levels of GST genes from Plutellaxylostella by…

Elsevier

Pesticide Biochemistry and Physiology

Volume 98, Issue 1, September 2010, Pages 121-127

Pesticide Biochemistry and Physiology

Characterization of acephate resistance in the diamondback moth Plutellaxylostella

Author links open overlay panelShojiSonodaChikakoIgaki

https://doi.org/10.1016/j.pestbp.2010.05.010Get rights and content

Abstract

Decreased acetylcholinesterase (AChE) sensitivity and metabolic detoxification mediated by

glutathione S-transferases (GSTs) were examined for their involvement in resistance to acephate in

the diamondback moth, Plutellaxylostella. The resistant strain showed 47.5-fold higher acephate

resistance than the susceptible strain had. However, the resistant strain was only 2.3-fold more

resistant to prothiofos than the susceptible strain. The resistant strain included insects having the

A298S and G324A mutations in AChE1, which are reportedly involved in prothiofos resistance in P.

xylostella, showing reduced AChE sensitivity to inhibition by methamidophos, suggesting that

decreased AChE1 sensitivity is one factor conferring acephate resistance. However, allele

frequencies at both mutation sites in the resistant strain were low (only 26%). These results suggest

that other factors such as GSTs are involved in acephate resistance. Expression of GST genes

available in P. xylostella to date was examined using the resistant and susceptible strains, revealing

no significant correlation between the expression and resistance levels.

See discussions, stats, and author profiles for this publication at: https://www.researchgate.net/publication/322163075

Detection of insecticide resistance and mechanisms of resistance in field

populations of Chrysoperla zastrowi sillemi (Neuroptera: Chrysopidae)

collected from different geographica...

Article  in  Journal of Biological Control · January 2017

DOI: 10.18311/jbc/2017/16333

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Sushil K. Jalali

National Bureau of Agricultural Insect Resources

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University of Bedfordshire

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61

Journal of Biological Control, 31(3): 61-68, 2017, DOI: 10.18311/jbc/2017/16333

Research Article

T. VENKATESAN1*, S. HELEN MAHIBA2, S. K. JALALI1, S. L. RAMYA1 and M. PRATIBHA11ICAR-National Bureau of Agricultural Insect Resources, P.B. No. 2491, H.A. Farm Post, Hebbal, Banaglore - 560024, Karnataka, India2Regional Seri cultural Research Station Central Silk Board, Veeranam Road, Allikuttai Post, Vaikkalapattarai, Salem - 636003, Tamil Nadu, India *Corresponding author E-mail: tvenkat12@gmail.com

Detection of insecticide resistance and mechanisms of resistance in field populations of Chrysoperla zastrowi sillemi (Neuroptera: Chrysopidae) collected from different geographical locations in India

(Article chronicle: Received: 22-06-2017; Revised: 21-09-2017; Accepted: 30-09-2017

ABSTRACT: The toxic effect of commonly used insecticides in cotton fields was studied on 9 populations of Chrysoperla zastrowi sil-lemi (Esben-Petersen), an important predator of sucking pests collected in India. The dose mortality bioassay against 3-days old larvae was determined using three insecticides viz., endosulfan, fenvalerate and acephate by topical bioassay method. Mechanism of resistance to the above mentioned insecticides were determined without and with three metabolic inhibitors (synergists), viz., piperonyl butoxide (PBO), S,S,S-tributyl-phosphorotrithioate (DEF) and diethyl maleate (DEM). Among the populations, resistant ratios (RR) of CZS-8 was significantly higher i.e. 50.36., 66.11 and 277.51-fold for endosulfan, fenvalerate and acephate, respectively compared to susceptible popu-lation (CZS-10). The CZS-8 was selected for synergism study it showed higher LC

50 values and resistance ratio for all three insecticides.

It showed 8.97-fold, 18.49-fold and 6.38-fold increase in synergism ratio for endosulfan indicating the resistance was strongly synergised by PBO, DEF and DEM. Similarly for fenvalerate, CZS-8 showed 8.69-fold and 3.63-fold significant increase in synergism ratio by DEF and DEM, respectively and for acephate, CZS-8 showed 54.82-fold, 150.87-fold and 113.52-fold significant increase in synergism ratio indicating that the resistance could be due to cytochrome p-450, esterase and glutathione s- transferase activity. The study indicated that the field population of C. z. sillemi developed resistance to different groups of insecticides. Among different geographical populations, CZS-8 collected from Sriganganagar, was recorded as most resistant.

KEY WORDS: Chrysoperla zastrowi sillemi, cytochrome p450, esterase, glutathione –S- transferase insecticide resistance

INTRODUCTION

The Common green lacewing, Chrysoperla zastrowi sillemi (Esben-Petersen) (Neuroptera: Chrysopidae), is an important biological control agent of sucking pests in dif-ferent agroecosystems (Symondron et al., 2002; Venkatesan et al., 2008; Henry et al., 2010). It has long been considered as a promising candidate for pest management programs worldwide due to its wide prey range and geographical dis-tribution, voracious larval feeding capacity and commer-cial availability (Medina et al., 2003; Pathan et al., 2010; Sayyed et al., 2010). Parasitoids and predators are highly susceptible to insecticides than their host insects [Croft and Brown, 1975), which make them difficult to establish in in-secticide sprayed field. Parasitoids and predators are known to develop resistance to insecticides in nature like their prey insects either by direct exposure or by consumption of prey insects treated with insecticides (Wu et al., 2004; Wu and

Miyata, 2005). However, resistance development is due to a combination of biological and ecological factors operating in the field (Venkatesan et al., 2009; Pathan et al., 2010). Compatibility of insecticide with biocontrol agents is im-portant as their application against the insect pests directly and indirectly determines the effectiveness of bioagents. In nature, populations of predators and insect pests always mutually co-exist often in a density-dependant association. Any adaptation of the insect pests with insecticide sprays is likely to be followed by the predator also to sustain them-selves in a given habitat.

In India, several chemical insecticides are used in-discriminately to control insect pests especially on cotton against sucking pests, which has led to resistance in many insect pests (Reddy and Rao, 1989; Kranthi et al., 2001). In a study conducted from 2007 to 2009, monocrotophos

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Detection of insecticide resistance and mechanisms of resistance in field populations of Chrysoperla zastrowi sillemi

resistance was documented in field populations of C. zas-trowi sillemi (Venkatesan et al., 2009). Chrysopid predators have been found resistant to insecticides in USA (Grafton and Hoy, 1985), Pakistan (Pathan et al., 2008; Sayyed et al., 2010), India (Venkatesan et al., 2009) & Canada (Pree et al., 2009). In a study, significantly higher fitness attributes viz., intrinsic rate, survival rate, doubling time and predation rate has been reported in organophosphate and pyrethroid resistant populations of C. carnea (Pathan et al., 2008) con-trary to general belief of genetic trade-off in such attributes in insects. However, information about the resistance level for different groups of insecticides and the mechanism(s) of resistance is important for successful augmentative releases of the resistant strain especially in the IPM of insect pests. Therefore, release of insecticide resistant predators would improve their survival in sprayed situations for potential use in augmentative biological control or integrated pest man-agement strategies in many crops. Further, such predators can play an effective role in delaying the development of re-sistance in pest populations and reduce the pest resurgence.

Metabolic enzymes play a significant role in detoxifi-cation of insecticides in insects (Motoyama, 1980). Mixed function oxidase, glutathione-s-transferase and esterase are involved in many insects in insecticide resistance mecha-nisms (Narahashi et al., 1995) due to their ability to de-toxify insecticides and other xenobiotics (Li et al., 2007). Many synergists such as piperonyl butoxide (PBO), diethyl maleate (DEM) and S,S,S-tributyl phosphoro trithioate (DEF) used at non-toxic doses are known to inhibit monox-ygenase, glutathione-s-transferase and esterase activities, respectively (Casida, 1970; Scott, 1990). However, work on insecticide resistance and mechanisms of resistance in chrysopid predators is very scanty and this is first kind of such study in India. Therefore, in the present study, based on the initial screening to representatives of three major groups of insecticides, namely endosulfan (cyclodiene), fenvalerate (synthetic pyrethroid) and acephate (organo-phosphate), a resistant strain of C. z. sillemi (CZS-8) was selected and effect of synergists (PBO, DEM & DEF), known to inhibit important detoxification routes, was inves-tigated to know the mechanisms of resistance in the resist-ant population of C. z. sillemi. Thus the study focuses on selection of an insecticide resistant predator C. z. sillemi which can be used as one of the important components in the pest management strategies especially under insecticide stressed crop conditions.

MATERIALS AND METHODS

Chrysoperla zastrowi sillemi populations

Nine populations of Chrysoperla z. sillemi (~100 lar-vae/adults) were collected in 2008-09 from heavily sprayed

cotton fields in 9 cotton growing districts in eight states, viz., Coimbatore (Tamil Nadu state) Anand (Gujarat state), Delhi state, Sirsa (Haryana state), Sriganganagar & Udaipur (Rajasthan state), Guntur (Andhra Pradesh state), Dhar-ward (Karnataka state) and Ludhiana (Punjab state). The pesticide use pattern was recorded from each collection site (Table 1). A laboratory population of C. z. sillemi originally maintained for the past 11 years at ICAR-National Bureau of Agricultural Insect Resources (NBAIR), Bangalore, In-dia, without exposing to insecticides for 125 generations was used in the study as susceptible population.

Laboratory rearing

Chrysoperla zastrowi sillemi populations were main-tained separately in the laboratory on UV exposed (15 watt for one h in UV hood) eggs of Corcyra cephalonica (Stain-ton). UV exposure of eggs was done in order to kill the em-bryo and facilitate the rearing of the Chrysoperla. Freshly emerged adults were transferred to oviposition chambers (14 cm x 9 cm) covered with muslin cloth. Cotton swabs dipped in water and the other with 50% honey, proteinex (Pfizer limited, Mumbai, India) (consisting of pre-digested protein enriched with vitamins, carbohydrates and miner-als), yeast and sucrose in the ratio of 1:1:1:1) and castor pollen grains was provided as adult feed and covered with perforated brown paper for egg laying. Eggs were collected at two-day intervals and kept for hatching with C. cepha-lonica eggs and the containers were covered with perforated brown paper. Freshly emerged larvae were kept individually in glass vials (4 x 2.5 cm) plugged with cotton and fed on Corcyra eggs. The rearing was done at 26±1ºC, 65±5% RH at a photoperiod of 14L: 10D in a plant growth chamber.

Insecticides

Commercial formulations of insecticides have been used for dosage mortality and synergism studies (Sayyed et al., 2010; Ahmad and Hollingwoth, 2004). Furthermore, field resistance has been always reported for commercial formulation of insecticides. Hence, the following formulat-ed insecticides were used for bioassays and also for syner-gism studies: endosulfan 35 EC (Excel Crop Care Limited, Mumbai), fenvalerate 20 EC (Aimco pesticides Limited, Mumbai, India) and acephate 75% SP (Jai Radhe Sales, Ahmedabad, Gujarat, India). All other chemicals were of analytical grade and purchased from Sigma-Aldrich (Bel-gium).

Dose mortality bioassays

Based on the field recommended dosage of endosul-fan (0.07%) (2.0 ml/litre), fenvalerate (0.04%) (0.2 ml/litre) and acephate (0.05%) (0.67 g/litre) in India, the following concentrations were used for the bioassay studies: endosul-

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fan (0,0625, 0.25, 0.5, 1.0, 2.0, 4.0, 8.0, 16.0, 32.0, 64.0, 74.0, 84.0, 94.0 ml/lit), fenvalerate (0.2, 0.4, 0.8, 1.6, 3.2, 6.4, 12.8, 25.6, 51.2, 61.2 ml/lit) and acephate (1.34, 2.68, 5.36, 10.72, 21.44, 42.88, 85.76, 171.52, 343.04, 686.08 gm/lit). These were applied on 3-d-old larvae of C. z. sil-lemi larvae in the weight range of 0.8 to 1.2 mg by using topical assays (Pathan et al., 2008). Each insecticide was tested with seven concentrations initially and as we did not get 50% mortality, the number of concentrations were fur-ther increased to 13, 10, 10 for endosulfan, acephate and fenvalerate, respectively. Each concentration was replicated thrice to determine the LC

50 value. The treated larvae were

provided with Corcyra eggs and were reared in a growth chamber at a temperature and RH as mentioned earlier. Untreated (control) larvae were treated with distilled water alone. At least 30 larvae were used for each concentration and in control. The mortality was recorded after 48 h and the larvae were considered dead if they did not move when prodded.

Synergism studies

CZS-8 population of C. Z. sillemi, which had highest LC

50 and resistant factor for all three groups of insecticides

was selected for synergism studies. For synergism assays, the synergist piperonyl butoxide (PBO; 0.5ul (0.5mg/100 ml) (90% purity), diethyl maleate (DEM: 0.5ul (0.5mg/100 ml) (97% purity) and S,S,S-tributyl phosphorotrithioate (DEF; 0.5ul (0.5mg/100 ml) (98% purity) were dissolved individually in a mixture of N,N,-dimethylformamide and tween-80 (3:1 by weight) and subsequently diluted with de-

ionised water (100-fold) [25]. Endosulfan @ 94.0 ml/liter, fenvalerate @ 61.2ml/liter and acephate @ 686.08 g/lit were mixed with water and a series of dilutions was made.

Data analysis

The results from all replicates for each insecticide were pooled and dose mortality regressions were computed by Probit analysis [Finney, 1952), using SPSS 16.0 software. Resistance ratio (RR) was calculated as LC

50 of the field

strain/LC50

of the susceptible strain. Synergism ratios and their confidence limits were calculated using the formula and statistics of dose ratios [Robertson and Preisler, 1992].

RESULTS AND DISCUSSION

Toxicity Bioassays

Among the 9 field and one laboratory populations of C. z. sillemi tested, Sriganganagar population (CZS-8) recorded maximum LC

50 for endosulfan (252.82 ml/lit.)

followed by the population from Delhi (CZS-5), Anand (CZS-4), Udaipur (CZS-9), Ludhiana (CZS-7) and Dhar-wad (CZS-2) and these were significantly different from all other populations (Table 2). The resistance ratio (RR) was highest (50.36-fold) in Sriganganagar (CZS-8) followed by Delhi (CZS-5) (26.02-fold), Anand (CZS-4) (21.46-fold) and Udaipur (CZS-9) population (15.4- fold). Non-overlap-ping test of significance indicated that between the popu-lations, there was significant difference the populations, however, all the resistant populations, viz., CZS-1 to CZS-9 were significantly different from susceptible population (P≤0.01).

Table 1. Insecticide usage at sampling sites of Chrysoperla zastrowi sillemi on cotton 2007-2009 cropping seasonsSl. No.

Collection site (District &State-wise)

Code No.

Collection period

Details of insecticides used and no. of sprays (year prior to collection)

Latitude Longitude

1 Coimbatore (Tamil Nadu)

CZS-1 April 2008 Triazophos, endosulfan, quinalphos, acephate 75% SP, fenitrothion 3 sprays/month)

11 º 00’N 77º 00’E

2 Dharwad (Karnataka)

CZS-2 Sep-2009 Imidaclopid 17.8 SL, thiomethaxam 70 WS, oxydemeton methyl 25 EC, dimethoate 30 EC & endosulfan, 35 EC (3-5 sprays/month)

15 º 27’N 75º 05’E

3 Guntur (Andhra Pradesh)

CZS-3 Dec.2008 Endosulfan,triazophos, profenphos, acephate 75 % SP, indoxocarb (4-5 times/month)

16 º 18’N 80º 29’E

4 Anand (Gujarat)

CZS-4 Nov. 2008 Fenvalerate 20 EC, endosulfan 35 EC, profenphos, spi-nosad 48 SC, acephate 75 % SP (3 sprays/month)

22 º 32’N 73º 00’E

5 Delhi CZS-5 Oct. 2008 Acephate 75 % WP, oxydemeton methyl 25 EC , dimethoate 30 EC (3 sprays/month)

28 º 38’N 77º 12’E

6 Sirsa (Haryana)

CZS-6 Oct. 2008 Acephate, triazophos, spinosad, indoxocarb, fenvalerate (4 sprays/month)

29 º 53’N 75º 020’E

7 Ludhiana (Punjab)

CZS-7 May 2009 Acephate, triazophos, spinosad, indoxocarb, fenvalerate (4 sprays/month)

30º 55’N 75º 54’E

8 SriGanganagar (Rajasthan)

CZS-8 Oct. 2008 oxydemeton methyl 25 EC , dimethoate 30 EC, acephate 75% SP, phosphamidon 85 WSC/ha (3-4 sprays/month)

29º 49’N 73º 50’E

9 Udaipur (Rajasthan)

CZS-9 Feb. 2009 Acephate 75 % WP, oxydemeton methyl 25 EC, dimethoate 30 EC, phosphamidon 85 WSC/ha (3-4 sprays/month)

27º 42’N 75º 33’E

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Detection of insecticide resistance and mechanisms of resistance in field populations of Chrysoperla zastrowi sillemi

In the test of significance by non-overlapping method, for fenvalerate, CZS-8 recorded high resistance (81.98 ml/litre) which was on par with Delhi, Ludhiana, Anand and Dharwad populations and were significantly different from remaining field populations. LC

50 of all the field popula-

tions were significantly different from susceptible popula-tion (P≤0.01). Resistance ratio (RR) for fenvalerate ranged from 9.12 to 66.11-fold in the 9 populations. The highest RR was recorded in CZS-8 (66.11-fold) followed by CZS-5 (38.89-fold), CZS-9 (25.91-fold), CZS-4 (21.64-fold) and

CZS-6 (17.94-fold) (Table 3).

Resistance to acephate was highest in Sriganganagar population (CZS-8) (535.60 g/litre) which was significantly at par with CZS-9, CZS-6, CZS-3 and CZS-1 and were sig-nificantly superior to rest of the populations (Table 4). The study showed that C. z. sillemi had cross resistance to dif-ferent groups of insecticides, viz., endosulfan, fenvalerate and acephate.

Table 3. Toxicity of fenvalerate to field collected and lab reared (susceptible) Chrysoperla zastrowi sillemiStrain n Slope + SE LC

50

g/lit or ml/lit95% FL χ2 Probability

P*RR

CZS-1 240 0.272+.214 11.32 b 7.02-19.21 3.36 0.971 9.12

CZS-2 225 0.640+0.529 17.52 a 8.08-57.84 1.39 0.966 14.12

CZS-3 211 1.19+0.933 14.93 b 8.92-22.48 1.23 1.000 12.04

CZS-4 223 0.66+0.517 26.84 a 16.93-58.12 1.59 0.991 21.64

CZS-5 210 2.35+1.48 48.23 a 36.46-99.20 0.127 0.988 38.89

CZS-6 215 0.455+0.359 22.25 b 13.57-44.77 5.37 0.865 17.94

CZS-7 210 0.620+0.448 24.26 a 12.73-54.70 3.26 0.860 19.56

CZS-8 215 1.112+0.704 81.98 a 48.72-758.39 0.847 0.932 66.11

CZS-9 230 2.004+1.26 32.14 b 18.4-46.07 1.48 0.993 25.91

CZS-10@ 212 0333+0.2 1.24 c 0.48-2.762 2.23 0.973

n = Number of larvae used in bioassay, including controls.RR= Resistance Ratio, calculated as LC

50 of field collected (or resistance) strain /LC

50 of susceptible

@= lab reared susceptible population Means within a column followed by different letters are significantly different (P

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VENKATESAN et al.

In India, insecticides are the most common means of controlling the pests by farmers and acephate, fenvaler-ate and endosulfan are the most widely used insecticides against sucking pests on cotton (Radika and Subbaratnam, 2006; Dhawan et al., 2009). Armes et al. (1994) reported that use of increasing number of insecticide brands, spu-rious insecticide use, lack of proper recommendations are the some of the reasons for the pest management problems in India. As a result insect pests developed resistance to different groups of insecticides which forces the farmers to go for increased number of insecticides to combat the pests. Kranthi et al. (2001) reported that many insect pests have developed resistance to these insecticides on cotton. Since the introduction of Bt cotton in India, frequency of insecticides applied against bollworms has come down drastically, however, sucking pests like aphids, whiteflies, thrips, mealybugs and leafhoppers are a serious bottleneck for successful cultivation of cotton and insecticides are in-creasingly applied to combat such pests. However, studies on development of insecticide resistance in natural enemies are very scanty. In this connection, insecticide resistant C. z. sillemi would be useful for the effective suppression of sucking pests as they can survive and multiply in sprayed situation.

Field strains of H. armigera exhibited widespread resistance to synthetic pyrethroid (cypermethrin) with 23–8022-fold resistances. Resistance to endosulfan (23-57-fold) and chlorpyriphos (4-82-fold) was low to high in H. armigera was observed. Besides, Spodoptera litura, Earias vitella and Bemisia tabaci from cotton filed developed mod-erate to high level of resistance to pyrethroid, organophos-phate and cyclodiene in India (Dhawan et al., 2009). Some

of the field collected populations of C. z. sillemi showed high resistance to acephate, fenvalerate and endosulfan in this study. This shows clearly that C. z. sillemi, which is the dominant predator found in cotton has developed resistance to different groups of insecticides along with insect pests in India. The enhanced resistance in Sriganganagar popula-tion (CZS-8) of C. z. sillemi correlates well with the greater use of insecticides in that region, particularly on cotton, where on average of 8-22 rounds of sprays of insecticides were used against a complex of insect pests [Kranthi et al., 2001]. The development of insecticide resistance in C. z. sillemi is primarily a result of the selection pressure exerted on sprayed populations increasing the frequency of resistant individuals which perhaps would have altered the genetic make-up of the organisms to survive and withstand higher doses of insecticides. Venkatesan et al. [2009] reported that out of the 9 field populations of C. z. sillemi, Sriganganagar (Rajasthan, India) population exhibited very high resistance to monocrotophos as compared to laboratory population. The study further supports our findings that the predator from Sriganganagar (CZS-8) developed resistance not only to monocrotophos but also to endosulfan, fenvalerate and acephate with RR increased to 50.36-fold, 66.11-fold and 277.51-fold, respectively, compared to susceptible. Sayyed et al. (2010) reported RRs of 47, 86, 137, 76 and 110 for deltamethrin, alphamethrin, lamdacyhalothrin, chlorpyrifos and profenofos for resistant C. carnea as compared to lab population in Pakistan, which was in conformity with our study. Natural tolerance to pyrethroid in C. carnea has been reported [Plapp and Bull, 1978). Further, Croft and Brown [1975] reported that natural enemies were more tolerant than their prey or host (67 of 92 cases) and predators were more tolerant than their prey (63 of 77 cases). Among the

Table 4. Toxicity of acephate to field collected and lab reared (susceptible) Chrysoperla zastrowi sillemiStrain n Slope + SE LC

50

g/lit or ml/lit95 % FL χ2 Probability

pRR

CZS-1 223 2.61+1.09 384.02 a 260.95-1565.43 0.958 1.000 198.97

CZS-2 210 0.792+0.617 5.02 c 3.49-7.07 1.033 0.984 2.60

CZS-3 230 2.47+0.952 501.63 a 326.93-1114.73 8.09 0.778 259.91

CZS-4 215 7.71+5.6 5.30 c 3.52-7.92 0.026 1.00 2.74

CZS-5 230 1.201+1.01 12.66 b 9.26-16.05 1.86 0.967 6.55

CZS-6 210 2.43+1.06 255.04 a 171.12-564.53 2.5 0.996 132.14

CZS-7 215 0.455+0.359 22.25 b 13.57-44.77 5.37 0.865 11.52

CZS-8 210 2.61+0.98 535.60 a 362.98-1041.18 1.84 0.985 277.51

CZS-9 218 1.93+0.782 317.88 a 192.49-585.41 .2.8 0.946 164.70

CZS-10@ 212 1.04+0.59 1.93 c 0.163-3.604 0.457 1.00

n = Number of larvae used in bioassay, including controls.RR= Resistance Ratio, calculated as LC

50 of field collected (or resistance) strain /LC

50 of susceptible

@= lab reared susceptible population Means within a column followed by different letters are significantly different (P

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Detection of insecticide resistance and mechanisms of resistance in field populations of Chrysoperla zastrowi sillemi

natural enemies, Amblyseius chilenensis was the first preda-tory mite found resistant to chemical pesticides (Kranthi et al., 2002). Similarly, insecticide resistance in different geographical populations of Chrysoperla carnea in Paki-stan was earlier reported [Pathan et al., 2008; Sayyed et al., 2010; Venkatesan et al., 2009). Hence it may correct that chrysopid predators in India and Pakistan is being increas-ingly developed resistance to insecticides especially from the cotton which could be due to heavy insecticidal sprays used to control the sucking pests especially for the newly introduced cotton mealybug Phenococus solenopsis.

Synergism studies

The population CZS-8 was selected for synergism studies based on their higher LC

50 and RR. PBO had dif-

ferent effects on the toxicity of endosulfan, fenvalerate and acephate (Table 5) to insecticide resistant (CZS-8) and sus-ceptible populations. PBO caused an 8.97-fold increase in

toxicity of endosulfan, 0.855-fold for fenvalerate and 54.32-fold for acephate. DEF caused an 18.49-fold increase in toxicity of endosulfan in CZS-8 population. The synergism of DEF on fenvalerate in CZS-8 population enhanced the toxicity by 8.69-fold and DEF showed obvious synergism. On acephate in the same strain, the synergism increased to 8.69-fold. The synergism of DEM on endosulfan enhanced the toxicity by 6.38-fold; 3.63-fold for endosulfan, fenva-lerate, respectively and 113.52-fold for acephate. Syner-gism was found to be very low when the effect of PBO was tested on resistance for fenvalerate in CZS-8. However, a synergistic effect could be detected in those populations when treated with DEF with endosulfan, fenvalerate and acephate. DEF produced a very high synergistic effect on acephate (SR ratio: 150.87) followed by endosulfan (SR ratio: 18.49) and fenvalerate (SR ratio: 8.69). This shows that DEF enhanced synergism in acephate, fenvalerate and endosulfan.

Table 5. Toxicity of endosulfan, fenvalerate and acephate with and without synergists to insecticide resist-ant and susceptible strains of Chrysoperla zastrowi sillemi

Population Treatment Slope ±SE LC50

(95% CL) SR

Lab Endosulfan 0.792+0.617 5.02 22-64.96 ----

+ PBO 0.239±0.220 3.202 1.23-6.64 1.57

+ DEF 0.268±0.232 4.85 1.22-11.16 1.03

+ DEM 0.268±0.243 2.35 0.32-5.56 2.1

CZS-8 Endosulfan 0.982+0.586 252.82 87.46-3.23 ---

+ PBO 1.26+0.875 28.180 19.89-42.02 8.97 a

+ DEF 1.146 + 0.997 13.67 9.99-19.13 18.49 a

+ DEM 0.502+ 0.298 39.62 25.06-63.33 6.38 a

Lab Fenvalerate 0.333+0.2 1.24 0.48-2.762 ---

+ PBO 0.204±0.182 1.041 0.26-5.296 1.19

+ DEF 1.088±0.281 1.152 0.106-2.88 1.07

+ DEM 0.254±0.233 1.23 0.01-4.86 1.00

CZS-8 Fenvalerate 1.112+0.704 81.98 48.72-758.39 ----

+ PBO 4.504+2.61 95.85 ---- 0.855

+ DEF 0.342+ 0.302 9.43 5.52-17.6 8.69 a

+ DEM 0.782+0.559 22.55 15.28-33.39 3.63 a

Lab Acephate 1.04+0.59 1.93 0.163-3.604 ---

+ PBO 0.198±0.181 1.101 0.006-4.818 1.75

+ DEF 0.239±0.191 1.554 0.64-2.9 1.24

+ DEM 0.214±0.153 1.492 0.026-27.05 1.29

CZS-8 Acephate 2.61+0.98 535.60 362.98-1041.18 ---

+ PBO 0.839+0.806 9.86 6.92-13.94 54.32 a

+ DEF 0.954+ 0.588 3.55 2.54-4.93 150.87 a

+ DEM 0.502+0.298 39.62 25.06-63.33 113.52 a

Synergism Ratio (SR)- LC50

of insecticide alone/LC50

of insecticide + synergist.Abbreviations: LC= Lethal Concentration expressed as gm/larva; FL= Fiducial limits; SE= Standard Errora There is significant synergism based on on-overlapping of the 95% CL’s of the LC

50 values between insecticide only and insecticide

after synergists treatment.

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VENKATESAN et al.

Pyrethroid resistance has been attributed to reduced neural sensitivity, enhanced metabolism and reduced pen-etration ratio in many insects [Oppenoorth, 1985; Zerba et al., 1987). Atkinson et al. (1991) reported that permethrin and cypermethrin resistance in a highly pyrethroid resist-ant strain of Blattella germanica was partially suppressed with PBO and DEF, thus suggesting the involvement of en-hanced metabolism as well as target site insensitivity in the mechanism of resistance. Picollo et al. [2000] reported that enhanced metabolism and synergism by enzyme inhibitor was involved in pyrethroid resistance in Pediculus capitis. The activation by midgut esterases from the tobacco horn-worm, Manduca sexta (L.) was inhibited by DEF (Kranthi et al., 2002). Sayyed et al. (2010) demonstrated that PBO reduced the LC

50 for deltamethrin (8 fold), alphamethrin

(3-fold) and lambdacyhalothrin (1.6-fold) in deltamethrin resistant strain of C. carnea which is in conformity with our present study.

In the current study, in case of resistance to acephate, PBO did decrease the resistance in CZS-8 population. This shows that the PBO block esterase activity which perhaps plays an important role in detoxification of acephate. Simi-larly, PBO had also been reported to inhibit resistance re-lated esterases in some insect species (Wing et al., 1998; Gunning et al., 1998; Gunning et al., 1999). DEF played a role in detoxification of endosulfan, fenvalerate and ace-phate in all the populations by increasing synergism ratio in the present investigation. This suggests that DEF could in-hibit monoxygenase, esterase and GST activities which are in accordance with earlier studies that it is not a completely specific inhibitor of esterase that it can also inhibit mon-oxygenase at high concentration (Young et al., 2005; Valles et al., 1997). Similarly, DEM also suppressed the toxicity of endosulfan, fenvalerate and acephate by increasing the synergism which indicates the activity of monoxygenase, esterase and GST. The combined evidence of in vitro and synergism bioassays indicate that the insecticide resistance in C. z. sillemi could be due to either enhanced esterase and or monooxygenase and GST activities.

Though synergism bioassays and in vitro enzyme as-says indicated that metabolic detoxification was an impor-tant resistance mechanism, the fact that full suppression of resistance was never achieved in any of the populations suggests that metabolic detoxification was probably one of the many mechanisms conferring insecticide resistance. Sayyed et al. (2010) reported that resistant natural enemies could be an alternative option to use them in concurrence with insecticides. They found that C. carnea developed cross resistance to pyrethroid and organophosphate com-pounds which is in accordance with our study.

Detoxification enzymes are similar in most of the insects including pests and natural enemies and the high esterase activity in lacewing larvae contributed to natural tolerance to pyrethroids (Dhawan et al., 2009). Bozsik et al. (2002) reported that C. carnea was tolerant to paraoxon (organophosphate group) due to higher activity of acetyl cholinesterase (AChE). Mixed-function oxidases (MFO) and hydrolysing esterases may be involved in detoxifica-tion of carbaryl resistant strain of C. carnea larvae. Fur-ther, Grafton & Hoy (1985) found that C. carnea possesses naturally high esterase enzyme levels that provide them natural resistance for pyrethroids. Further, monoxygenase-mediated resistance to pyrethroids was found in C. carnea (Pree et al., 1989) which are in conformity with our study. Insecticide resistant selected C. carnea may tolerate insec-ticide pressure in the field conditions (Sayyed et al., 2010). The study revealed the selection of insecticide resistant C. z. sillemi which can be used in the IPM programs. Sayyed et al. (2010) opined that release of insecticide resistant C. carnea will survive for the field dosage of pesticides and also inherit all genes involved in insecticide resistance to subsequent generations.

The study clearly showed that the field populations of Chrysoperla zastrowi sillemi from cotton developed resist-ance for insecticides belonging to pyrethroids, organophos-phate and cyclodiene. Among the resistant populations, CZS-8 had been found to have greater RRs to different insecticides, hence may be considered for the field evalu-ation. This is the first kind of such study in India. Mass production and release of such resistant predator would im-prove their survival in sprayed situations for potential use in augmentative biological control or integrated pest man-agement strategies in not only on cotton but also on other crops. Further, such predators can play an effective role in suppressing the insecticide resistant pest populations and resurgence of secondary pests.

ACKNOWLEDGEMENTS

The authors are also grateful to the Director, ICAR-NBAIR, Bangalore, India for providing necessary facilities for conducting the study.

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US Environmental Protection Agency Office of Pesticide Programs

Reregistration Eligibility Decision for Acephate

Combined PDF document consists of the following:

• Finalization of Interim Reregistration Eligibility Decisions (IREDs) and Interim Tolerance Reassessment and Risk Management Decisions (TREDs) for the Organophosphate Pesticides, and Completion of the Tolerance Reassessment and Reregistration Eligibility Process for the Organophosphate Pesticides (July 31, 2006)

• Acephate IRED

When EPA concluded the organophosphate (OP) cumulative risk assessment in July 2006, all tolerance reassessment and reregistration eligibility decisions for individual OP pesticides were considered complete. OP Interim Reregistration Eligibility Decisions (IREDs), therefore, are considered completed REDs. OP tolerance reassessment decisions (TREDs) also are considered completed.

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY WASHINGTON D.C., 20460

OFFICE OF

PREVENTION, PESTICIDES AND TOXIC

SUBSTANCES

MEMORANDUM

DATE: July 31, 2006

SUBJECT: Finalization of Interim Reregistration Eligibility Decisions (IREDs) and Interim Tolerance Reassessment and Risk Management Decisions (TREDs) for the Organophosphate Pesticides, and Completion of the Tolerance Reassessment and Reregistration Eligibility Process for the Organophosphate Pesticides

FROM: Debra Edwards, Director Special Review and Reregistration Division Office of Pesticide Programs

TO: Jim Jones, Director Office of Pesticide Programs

As you know, EPA has completed its assessment of the cumulative risks from the organophosphate (OP) class of pesticides as required by the Food Quality Protection Act of 1996. In addition, the individual OPs have also been subject to review through the individual-chemical review process. The Agency’s review of individual OPs has resulted in the issuance of Interim Reregistration Eligibility Decisions (IREDs) for 22 OPs, interim Tolerance Reassessment and Risk Management Decisions (TREDs) for 8 OPs, and a Reregistration Eligibility Decision (RED) for one OP, malathion.1 These 31 OPs are listed in Appendix A.

EPA has concluded, after completing its assessment of the cumulative risks associated with exposures to all of the OPs, that:

(1) the pesticides covered by the IREDs that were pending the results of the OP cumulative assessment (listed in Attachment A) are indeed eligible for reregistration; and

1 Malathion is included in the OP cumulative assessment. However, the Agency has issued a RED for malathion, rather than an IRED, because the decision was signed on the same day as the completion of the OP cumulative assessment.

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(2) the pesticide tolerances covered by the IREDs and TREDs that were pending the results of the OP cumulative assessment (listed in Attachment A) meet the safety standard under Section 408(b)(2) of the FFDCA.

Thus, with regard to the OPs, EPA has fulfilled its obligations as to FFDCA tolerance reassessment and FIFRA reregistration, other than product-specific reregistration.

The Special Review and Reregistration Division will be issuing data call-in notices for confirmatory data on two OPs, methidathion and phorate, for the reasons described in detail in the OP cumulative assessment. The specific studies that will be required are:

− 28-day repeated-dose toxicity study with methidathion oxon; and − Drinking water monitoring study for phorate, phorate sulfoxide, and phorate sulfone

in both source water (at the intake) and treated water for five community water systems in Palm Beach County, Florida and two near Lake Okechobee, Florida.

The cumulative risk assessment and supporting documents are available on the Agency’s website at www.epa.gov/pesticides/cumulative and in the docket (EPA-HQ-OPP-2006-0618).

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http://www.epa.gov/pesticides/cumulative

Attachment A: Organophosphates included in the OP Cumulative Assessment

Chemical Decision Document Status Acephate IRED IRED completed 9/2001 Azinphos-methyl (AZM) IRED IRED completed 10/2001 Bensulide IRED IRED completed 9/2000 Cadusafos TRED TRED completed 9/2000 Chlorethoxyphos TRED TRED completed 9/2000 Chlorpyrifos IRED IRED completed 9/2001 Coumaphos TRED TRED completed 2/2000 DDVP (Dichlorvos) IRED IRED completed 6/2006 Diazinon IRED IRED completed 7/2002 Dicrotophos IRED IRED completed 4/2002 Dimethoate IRED IRED completed 6/2006 Disulfoton IRED IRED completed 3/2002

Ethoprop IRED IRED completed 9/2001 IRED addendum completed 2/2006 Fenitrothion TRED TRED completed 10/2000 Malathion RED RED completed 8/2006 Methamidophos IRED IRED completed 4/2002 Methidathion IRED IRED completed 4/2002 Methyl Parathion IRED IRED completed 5/2003 Naled IRED IRED completed 1/2002 Oxydemeton-methyl IRED IRED completed 8/2002 Phorate IRED IRED completed 3/2001 Phosalone TRED TRED completed 1/2001 Phosmet IRED IRED completed 10/2001 Phostebupirim TRED TRED completed 12/2000 Pirimiphos-methyl IRED IRED completed 6/2001 Profenofos IRED IRED completed 9/2000 Propetamphos IRED IRED completed 12/2000 Terbufos IRED IRED completed 9/2001 Tetrachlorvinphos TRED TRED completed 12/2002 Tribufos IRED IRED completed 12/2000 Trichlorfon TRED TRED completed 9/2001

Page 3 of 3

United States Prevention, Pesticides EPA 738-R-01-013 Environmental Protection and Toxic Substances September 2001

Agency (7508C)

Reregistration Eligibility Decision for Acephate

United States Prevention, Pesticides EPA 738-F-01-013 Environmental Protection and Toxic Substances September 2001 Agency (7508C)

Acephate Facts EPA has assessed the risks of acephate and reached an Interim Reregistration Eligibility

Decision (IRED) for this organophosphate (OP) pesticide. Provided that risk mitigation measures are adopted, acephate fits into its own “risk cup”-- its individual, aggregate risks are within acceptable levels. Acephate also is eligible for reregistration, pending a full reassessment of the cumulative risk from all OPs.

Acephate residues in food and drinking water do not pose risk concerns, and by reducing exposure in homes and through residential lawns, acephate fits into its own “risk cup.” EPA made this determination after the registrants agreed to drop indoor residential uses and certain turf uses. With other mitigation measures, acephate’s worker and ecological risks also will be below levels of concern for reregistration.

EPA’s next step under the Food Quality Protection Act (FQPA) is to consider risks from cumulative exposure to all the OP pesticides, which share a common mechanism of toxicity. The interim decision on acephate cannot be considered final until the cumulative risk has been considered. Further risk mitigation may be warranted at that time.

EPA is reviewing the OP pesticides to determine whether they meet current health and safety standards. Older OPs need decisions about their eligibility for reregistration under FIFRA. OPs with residues in food, drinking water, and other nonoccupational exposures also must be reassessed to make sure they meet the new FQPA safety standard.

The OP Pilot Public Participation Process

The organophosphates are a group of related pesticides that affect the functioning of the nervous system. They are among EPA’s highest priority for review under the Food Quality Protection Act.

EPA is encouraging the public to participate in the review of the OP pesticides. Through a six-phased pilot public participation process, the Agency is releasing for review and comment its preliminary and revised scientific risk assessments for individual OPs. (Please contact the OP Docket, telephone 703-305-5805, or see EPA’s web site, www.epa.gov/pesticides/op .)

EPA is exchanging information with stakeholders and the public about the OPs, their uses, and risks through Technical Briefings, stakeholder meetings, and other fora. USDA is coordinating input from growers and other OP pesticide users.

Based on current information from interested stakeholders and the public, EPA is making interim risk management decisions for individual OP pesticides, and will make final decisions after the cumulative risk from all OPs has been considered.

The acephate interim decision was made through the OP pilot public participation process, a process that increases transparency and maximizes stakeholder involvement in EPA’s development of risk assessments and risk management decisions. EPA worked extensively with affected parties to reach the decisions presented in this interim decision document that concludes the OP pilot process for acephate.

Uses

• Acephate is an organophosphate insecticide currently registered for use on a variety of field, fruit, and vegetable crops (e.g., cotton, tobacco, cranberries, mint); in food handling establishments; on ornamental plants both in greenhouses and outdoors (e.g., nonbearing fruit trees, Christmas trees, and cut flowers); and in and around the home.

• Annual domestic use is approximately 4 to 5 million pounds of active ingredient per year.

Health Effects

• Acephate can cause cholinesterase inhibition in humans; that is, it can overstimulate the nervous system causing nausea, dizziness, confusion, and at very high exposures (e.g., accidents or major spills), respiratory paralysis and death.

Risks

• Dietary exposures to acephate from eating food crops treated with acephate are below the level of concern for the entire U.S. population, including infants and children. Drinking water is not a significant source of acephate exposure. However, people in the U.S. may be exposed to amounts of the acephate degradate methamidophos through food and drinking water as a result of acephate use. This exposure will be more fully addressed in the methamidophos IRED.

• EPA found risks are of concern for homeowners and children entering homes and lawn areas treated with acephate (excluding golf courses and spot or mound treatments for ant control).

• For agricultural and turf/Pest Control Operator (PCO) uses of acephate, several mixer/loader/applicator risk scenarios currently exceed the Agency’s level of concern. In addition, there are postapplication risks from the use of acephate in cut flowers.

• Ecological risks are also of concern to the Agency. Acephate and its degradate methamidophos are highly toxic to honey bees and beneficial predatory insects on an acute contact basis. Acute and chronic risks to birds and chronic risk to mammals are also high.

2

Risk Mitigation

Dietary Risk

No mitigation is necessary at this time for any dietary exposure to acephate. The acute and chronic dietary risks from acephate do not exceed the Agency’s level of concern.

However, the Agency reserves the right to require further acephate mitigation to address risks from methamidophos residues resulting from acephate uses. Any additional mitigation measures will be addressed when the methamidophos interim RED is completed.

Occupational Risk

In order to mitigate occupational risks, the following risk mitigation measures are necessary:

• Formulate all soluble powder formulations into water soluble bags, except for soluble powders sold for fire ant, harvester ant, or hopper box seed treatment uses.

• Limit the 1 pound active ingredient per acre (lb ai/A) cotton aerial application rate to cotton grown in California and Arizona; reduce the maximum aerial application rate for cotton to 0.75 ai/A for all other areas of the United States.

• Delete aerial application to turf. • Require enclosed cockpits and mechanical flagging for all aerial applications. • Reduce maximum sod farm and golf course turf application rates (non-granular formulations)

to 3 lb ai/A and 4 lb ai/A, respectively. • Reduce maximum application rates for greenhouse floral and foliage plant crops, and outdoor

floral and ground covers to 1 lb ai per 100 gallons water (not to exceed 0.75 lb ai/A for cut flowers and 1.0 lb ai/A for other ornamentals).

• Delete the application of acephate by low pressure handwand to treat trees, shrubs, and outdoor flora; for the control of wasps; and for perimeter treatment by PCOs.

• Delete the use of granular formulations to be applied by belly grinder, shaker can, or by hand to trees, shrubs, and 12" pots.

• Add personal protective equipment to end use product labels for workers who mix and load, and/or apply acephate.

Residential Risk

In order to mitigate residential postapplication risk, the following risk mitigation measures are necessary:

• Delete residential indoor uses. • Delete all turfgrass uses (except golf course, sod farm, and spot or mound treatment for ant

control). • Establish a 3 day pre-harvest interval (PHI) for the harvesting of sod.

3

Ecological Risk

The Agency has determined that the following mitigation measures are needed to address ecological risk concerns:

• Establish minimum spray intervals for all agricultural crops of 3 days for application rates up to 0.5 lb ai/A and of 7 days for application rates greater than 0.5 lb ai/A.

• Require labeling to protect honeybees. • Require labeling to reduce the potential for spray drift.

In addition, the measures to reduce occupational and residential risk will also reduce environmental loading and the potential impact to non-target organisms.

Next Steps

• Numerous opportunities for public comment were offered as this decision was being developed. The acephate IRED therefore is issued in final (see www.epa.gov/pesticides/reregistration/status.htm or www.epa.gov/pesticides/op ), without a formal public comment period. The docket remains open, however, and any comments submitted in the future will be placed in this public docket.

• In addition, further mitigation of acephate uses may be necessary to reduce risks from methamidophos residues that result from acephate applications. Once the methamidophos IRED is complete, the Agency will determine whether the methamidophos exposure resulting from acephate use poses risk concerns. Any potential further mitigation will be discussed at the time the methamidophos interim RED is released.

• When the cumulative risk assessment for all organophosphate pesticides is completed, EPA will issue its final tolerance reassessment decision for acephate and may request further risk mitigation measures. The Agency will revoke 3 tolerances and lower 4 tolerances for acephate now. Reassessment of 14 tolerances will be made once additional residue data on cotton gin byproducts have been reviewed. For all OPs, raising and/or establishing tolerances will be considered once a cumulative assessment is completed.

4

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY WASHINGTON, D.C. 20460

OFFICE OF PREVENTION, PESTICIDES AND TOXIC SUBSTANCES

CERTIFIED MAIL

Dear Registrants:

This is to inform you that the Environmental Protection Agency (hereafter referred to as EPA or the Agency) has completed its review of the available data and public comments received related to the preliminary and revised risk assessments for the organophosphate pesticide acephate. The public comment period on the revised risk assessment phase of the reregistration process is closed. Based on comments received during the public comment period and additional data received from the registrants, the Agency revised the human health and environmental effects risk assessments and made them available to the public on February 22, 2000. Additionally, the Agency held a Technical Briefing on February 2, 2000, where the results of the revised human health and environmental effects risk assessments were presented to the general public. This Technical Briefing concluded Phase 4 of the OP Public Participation Pilot Process developed by the Tolerance Reassessment Advisory Committee (TRAC), and initiated Phase 5 of that process. During Phase 5, all interested parties were invited to participate and provide comments and suggestions on ways the Agency might mitigate the estimated risks presented in the revised risk assessments. This public participation and comment period commenced on February 22, 2000, and closed on April 24, 2000.

Based on its review, EPA has identified risk mitigation measures that the Agency believes are necessary to address the human health and environmental risks associated with the current use of acephate. The EPA is now publishing its interim decision on the reregistration eligibility of and risk management decision for the current uses of acephate and associated human health and environmental risks. The reregistration eligibility and tolerance reassessment decisions for acephate will be finalized once the cumulative assessment for all of the organophosphate pesticides is complete. The enclosed “Interim Reregistration Eligibility Decision for Acephate” was approved on September 28, 2001, and contains the Agency’s decision on the individual chemical acephate.

A Notice of Availability for this Interim Reregistration Eligibility Decision (interim RED) is being published in the Federal Register. To obtain a copy of the interim RED document, please contact the OPP Public Regulatory Docket (7502C), US EPA, Ariel Rios Building, 1200 Pennsylvania Avenue NW, Washington, DC 20460, telephone (703) 305-5805. Electronic copies of the interim RED and all supporting documents are available on the Internet. See

http:www.epa.gov/pesticides/op.

The interim RED is based on the updated technical information found in the acephate public docket. The docket includes background information and comments on the Agency’s preliminary risk assessments; the Agency’s February 3, 2000, revised risk assessment for acephate; addenda to the occupational and residential risk assessments (September 15, 2000 and February 13, 2001); a revised surface water assessment (March 8, 2000); and a document summarizing the Agency’s Response to Comments. The Response to Comments document addresses corrections to the preliminary risk assessments submitted by chemical registrants and responds to comments submitted by the general public and stakeholders during the comment period on the risk assessment. The docket also includes comments on the revised risk assessment, and any risk mitigation proposals submitted during Phase 5. For acephate, a proposal was submitted by Valent U.S.A. Corporation (Valent), a technical registrant. All other technical registrants have agreed to the mitigation measures proposed. Comments on mitigation or mitigation suggestions were submitted by growers, agricultural extension agents, environmental organizations, university scientists, and various other organizations.

This document and the process used to develop it are the result of a pilot process to facilitate greater public involvement and participation in the reregistration and/or tolerance reassessment decisions for these pesticides. As part of the Agency’s effort to involve the public in the implementation of the Food